Conventional protocols for adult and pediatric tuberculosis (TB) diagnosis and treatment monitoring rely heavily on time-consuming bacterial culture or unquantifiable DNA detection methods for the presence of small numbers of bacteria. For pediatric TB, diagnosis and treatment are particularly difficult because current clinical protocols demand much from these young patients. Although they comprise a small percentage of the health care caseload, children who are co-infected with human immunodeficiency virus (HIV) and TB represent one of the most vulnerable groups with one of the highest mortality rates. We choose this challenging cohort (provided by a NIAID-sponsored clinical trial) to push the boundaries of what our technology platform can do. In order to address the current limitations in clinical management of pediatric TB, we have developed a rapid blood-based diagnostic method independent of mycobacterial isolation to quantify the low molecular- weight Mycobacterium tuberculosis (Mtb) antigens (CFP-10 and ESAT-6). Characteristics of ESAT-6 and CFP- 10 make them ideal biomarkers for active TB diagnosis and candidates for TB vaccine development. Our strategy combines energy mediating porous silicon nanodisks (referred to as pSiND), functionalized with customized antibodies highly specific to Mtb antigen peptides, and high-throughput mass spectrometry (NanoDisk-MS) for dual enhancement of sensitivity and specificity. We evaluated our platform with 292 adult and 102 pediatric patients and controls chosen from five highly relevant cohorts (active TB, HIV/TB co- infection, pediatric TB, latent TB, and non-TB mycobacterial infection), provided by multiple institutes worldwide. Sensitivities and specificities of adults (90.7% / 97.7%) and children (88.2% / 100%) were achieved in active TB identification. Absolute quantification of circulating antigens was informative in detecting treatment response four days after anti-mycobacterial initiation. Just as important, we could render accurate diagnoses within one hour of sample-to-answer processing rather than wait the typical 4-6 weeks. In this proposal, we aim to: 1) design and develop the scale-up nanodisk microfabrication protocol with FDA compliant in cGMP facilities; 2) conduct an extensive clinical validation of pSiND-MS using samples from a large cohort of children with HIV/TB; 3) determine effectiveness of our approach for rapid evaluation of treatment efficacy; and 4) optimize development and clinical validation of a portable pSiDN-miniMS system for identification and quantification of CFP-10 and ESAT-6. The pSiND-MS technology platform has an added advantage in that high-throughput and accurate mass spectrometry has become a virtually essential technology for clinical diagnosis in many parts of the world. The miniaturized and easy-to-use MS system at a shoebox size for point-of-care applications is aimed at serving patients in resource-limited areas. Achievement of all of our aims will significantly alter clinica management strategies for global TB control, and potentially improve diagnosis of other infectious diseases by quantifying circulating antigens of pathogenic organisms.